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Sensitizing ER-Negative Breast Cancer Cells to Tamoxifen-Induced Apoptosis by a Novel PDK-1 Inhibitor
Sensitizing ER-Negative Breast Cancer Cells to Tamoxifen-Induced Apoptosis by a Novel PDK-1 Inhibitor. Tamoxifen exhibits both estrogen receptor (ER)- and non-ER-mediated antiproliferative effects in breast cancer cells, though the latter occurs at therapeutically unattainable concentrations in vitro. OSU-03012, a novel celecoxib-derived phosphoinositide-dependent kinase-1 (PDK-1) inhibitor, potentiates the apoptotic effect of tamoxifen in both ER-positive and –negative breast cancer cells. With 5 µM OSU-03012, the IC50 values for tamoxifen-induced apoptosis in MCF-7 and MDA-MB-231 cells were 1.5 and 2.5 µM, respectively, vis-à-vis 15 µM for tamoxifen alone. This effect, however, was not noted with the pure antiestrogen ICI 182780, suggesting that ER does not play a role in this synergy. We rationalize that the mechanism underlying this synergistic effect is twofold: 1) blockade of Akt activation through PDK-1 inhibition synergizes with tamoxifen’s non-ER targets in facilitating apoptosis signaling, and 2) tamoxifen stimulates Akt activation. Thus, OSU-03012 renders cancer cells more susceptible to non-ER-mediated apoptosis by abrogating tamoxifen-induced Akt activation. These findings provide a rationale for the hypothesis that the synergistic interactions between Akt inhibition and tamoxifen’s non-ER targets can be clinically exploited to extend tamoxifen therapy to ER-negative breast cancer. The specific aims include: 1) in vivo validation of the synergy between OSU-03012 and tamoxifen in MCF-7 and MDA-MB-231 xenograft tumor models, and 2) dose optimization of the combination therapy in an MDA-MB-231 xenograft model. Aim 1 will demonstrate that ER does not play a role in the ability of OSU-03012 to enhance the in vivo antiproliferative effects of tamoxifen, and that Akt downregulation underlies the synergistic effect of OSU-03012 on tamoxifen-induced apoptosis. The effects of tamoxifen and ICI 182780, alone or with OSU-03012, on xenograft tumor growth will be assessed by analyzing proliferation and apoptotic indices, and phospho-Akt levels. Characterization of these in vivo biomarkers will provide correlations for the activities and mechanisms established in our in vitro studies that may verify the in vivo mechanism of synergistic action. Aim 2 will compare the effect of different doses of tamoxifen and OSU-03012 on MDA-MB-231 tumor growth to select an optimal combination for pharmacokinetic analysis. We expect the proposed studies to yield important data that can be translated to meet an urgent need in the treatment of ER-negative breast cancer.
Sensitizing ER-Negative Breast Cancer Cells to Tamoxifen-Induced Apoptosis by a Novel PDK-1 Inhibitor. There is an urgency to develop novel therapeutic strategies for estrogen receptor (ER) negative breast cancers as they do not benefit from hormonal therapies. We have developed a novel inhibitor of phosphoinositide-dependent kinase-1 (PDK-1), OSU-03012, which is currently undergoing preclinical evaluations at NCI. PDK-1 activates Akt-mediated survival pathways that underlie resistance to tamoxifen and chemotherapy in breast cancer. We tested OSU-03012 in combination with tamoxifen in ER-positive (MCF-7) and -negative (MDA-MB-231) breast cancer cells. The results show that OSU-03012 greatly enhanced the apoptotic effects of tamoxifen irrespective of the ER status. With 5 µM OSU-03012, the concentrations of tamoxifen to inhibit growth by 50% were 1.5 and 2.5 µM in MCF-7 and MDA-MB-231 cells, respectively, vis-à-vis 15 µM for tamoxifen alone. Tamoxifen concentrations of 1-3 µM are easily achievable in humans. This effect, however, was not noted in the combination of OSU-03012 and ICI 182780, a pure antiestrogen. We also found that tamoxifen caused an increase in activated Akt, but this Akt activation did not occur when OSU-03012 was present. These findings suggest that the ER may not play a role in the effect of OSU-03012 on tamoxifen-induced apoptosis, and that the inhibition of Akt activation is central to this synergy. We hypothesize that the synergy between OSU-03012 and tamoxifen may be clinically useful for the treatment of ER-negative breast cancer. This proposal seeks to extend our preliminary observations by testing OSU-03012 in combination with tamoxifen or ICI 182780 in MCF-7 and MDA-MB-231 xenograft models. Aim 1 will evaluate tumors from drug-treated animals for increased antiproliferative effects and changes in activated Akt. If our hypothesis is correct, the combined use of OSU-03012 and tamoxifen, but not ICI 182720, will enhance growth inhibition, increase apoptosis, and decrease activated Akt in ER negative tumors. Aim 2 will vary the doses of OSU-03012 and tamoxifen to define the most effective and least toxic dose. These animal experiments are crucial to bring the combined regimen into the clinic for ER-negative metastatic breast cancer. If this combination proves useful and establishes an important proof of principle, this will represent the first oral non-chemotherapy treatment for ER negative breast cancer, and will be a major advance in the treatment of early stage ER negative breast cancer, and possibly in breast cancer prevention.